Epidemiological studies have demonstrated that nonsteroidal antiinflammatory drugs (NSAIDs) can reduce the incidence of colon cancer. Since the well-documented pharmacological action of aspirin and other NSAIDs is inhibition of cyclooxygenase [COX, the rate limiting enzyme in prostaglandin (PG) biosynthesis], it can be inferred that the beneficial effect of NSAIDs may be mediated through the inhibition of PG biosynthesis. However, several lines of experimental observations imply that the beneficial effects of NSAIDs may be mediated through both COX-dependent and COX-independent pathways. Thus, Specific Aims are: 1) To determine whether the overexpression of COX in colon cancer cell lines or normal intestinal epithelial cells by transfecting with constitutively expressed COX-1 or inducible COX-2 cDNA, results in changes in tumorigenic phenotypes, and whether inhibiting COX by NSAIDs abrogates the changes in vitro (cells in culture) and in vivo (growth of transplanted tumors to athymic nude mice). Results from these studies will establish or negate a direct link of COX to tumor cell growth. 2) To determine whether cyclooxygenase-independent effects of NSAIDs are mediated through mitogen-activated protein kinase (MAPK), NFkappaB or peroxisome proliferator-activated receptor (PPAR) signaling pathways. Recent evidence suggests that NSAIDs modulate MAPK, NFkappaB and PPAR signaling pathways. Thus, these studies are aimed to identify COX-independent signaling pathways through which NSAIDs suppress tumorigenesis. 3) To identify differentially expresssed genes in the colon cancer line and normal intestinal epithelial cells overexpressing COX-2 by subtractive hybridization and Serial Analysis of Gene Expression methods. Identifying up-or-down-regulated genes caused by the overexpression of COX-2 or NSAID treatment in the cancer cells will provide a clue as to how increased prostaglandin production can lead to changes in tumorigenic phenotypes, or a clue to identifying the cellular targets (other than COX) of the NSAID actions. 4) To determine whether prostaglandin receptors that activate adenylate cyclase are differentially expressed in the colon cancer cells as compared with normal cells and to identify downstream signaling pathways of prostaglandin E receptor (Ep2). Understanding the roles of prostaglandins and their signaling pathways in cancer cells could help explore new treatments and/or preventive strategies for colon cancer and perhaps other cancers using pharmacological agents and/or dietary means that modulate prostaglandin biosynthesis.